High-molecular weight impurity screening by size-exclusion chromatography on a reversed-phase column

• Published in: Journal of pharmaceutical and biomedical analysis 2021-03, Vol.196, p.113908-113908, Article 113908
• Main Authors: Lin, Ziqing, Ye, Yun K., Ling, Melissa, Shackman, Jonathan G., Ileka, Kevin M., Raglione, Thomas V.
• Format: Article
• Language: English
• Subjects: C18 stationary phase; Pharmaceutical high-molecular weight impurity; Screening platform; Size-exclusion chromatography
• ISSN: 0731-7085; 1873-264X
• DOI: 10.1016/j.jpba.2021.113908

[Display omitted] •High-molecular weight impurity screening established on a single C18 column.•Nine mobile phase conditions to cover various polymeric impurities.•Real polymeric impurities monitored with a quantitative limit better than 0.1 % w/w. Monitoring polymerization events leading to the discovery of new high-molecular weight (MW) impurities is challenging during chemical syntheses of active pharmaceutical ingredients. Employing reversed-phase chromatography (RPC) stationary phases (SPs) in size-exclusion chromatography (SEC) mode could be a potential solution given their high efficiency, sensitivity, and extensive solvent compatibility. However, there is a lack of generalized means for trace polymeric impurities across a wide range of physicochemical properties. Herein, we developed a SEC-based approach with a C18 SP for screening such high-MW impurities. Seven polymer standards presenting a variety of functional groups, consisting of hydrophobic, heterocyclic, ionic, and neutral hydrophilic moieties, were utilized as model impurities to establish the screening conditions. Nine mobile phases (tetrahydrofuran-based, buffered methanol, and buffered acetonitrile) were proposed to cover all model polymers and a majority of potential high-MW impurities in small molecule chemical syntheses. The established screening system demonstrated a linearity of 0.05–1.0 % w/w (R2>0.99) for the selected model impurities with proper elution conditions. Two real high-MW impurities, BMT-041910 (polymeric degradation) and poly(phenyl thiirane) (by-product polymerization), were identified from the proposed high-MW impurity screening. The successful conditions yielded a quantitative limit better than 0.1 % w/w in both cases. We believe the developed screening platform is applicable to the analysis of a wide variety of unknown high-MW impurities of low abundance potentially generated during drug substance development.